Hypoxia-induced mitogenic factor (HIMF), also called “found in inflammatory zone 1” (FIZZ1) or resistin like molecule alpha (RELMα) or resistin, is induced in the lungs of experimental models of hypoxia-induced pulmonary hypertension (1, 2). We have shown that HIMF is expressed in the remodeling of hyperplastic vascular smooth muscle in vivo and can stimulate pulmonary microvascular mitogenesis in vitro. Also, we have demonstrated that HIMF possesses angiogenic and vasoconstrictive properties. In addition, HIMF increases pulmonary artery pressure and pulmonary vascular resistance more potently than endothelin, angiotensin or serotonin (1). Not only was HIMF expression increased in the lung vasculature, it was markedly up-regulated in hypertrophic, hyperplastic bronchial epithelium during allergic pulmonary inflammation in mouse models of acute pulmonary inflammation (3). A recent report has also demonstrated HIMF to be expressed in the lymph nodes (4), with the highest expression in B cells and macrophages.
The FIZZ/resistin gene family has been implicated in a variety of human diseases, but their binding partners/receptors until now had not been identified. Resistin which is one of two human analogs of HIMF, has recently been implicated in human vascular disease related to diabetes and to obesity (5). Resistin is greatly up-regulated in obesity and after insulin treatment, thus providing a potential molecular link between obesity and diabetes. It may function as a regulator of glucose homeostasis and an antagonist to insulin action (6). XCP1/FIZZ2 was found to be a chemotactic factor to myeloid cells from C/EBP-epsilon-null mice and is able to interact directly with alpha-defensin (7). There are three members of the FIZZ/resistin family that are encoded by two different genes in human, and five members of the FIZZ/resistin family that are derived from four genes in mouse. Receptors have not yet been identified for any of these physiologically and pathologically important rodent or human isoforms, leaving our functional understanding of this family of proteins incomplete.
Increasing evidence indicates that bone marrow-derived endothelial progenitor cells (EPCs) or circulating hematopoietic stem cells play an important role in postnatal neovascularization of adult ischemic tissues and injured tissues (8-11). Endogenous stimuli like tissue ischemia and exogenous cytokine therapy mobilize EPCs and thereby contribute to the neovascularization of ischemic and injured tissues (12). A number of chemokines have been reported to act as chemotactic and angiostatic molecules in inducing endothelial cell migration and regulating angiogenesis (13,14). In inflammatory diseases, angiogenesis and inflammatory disorders are two inter-related processes regulated by chemokines. XCP1, another member of FIZZ/resistin family sharing 70% identity in amino acid sequence with HIMF, has been reported to be a secreted protein that is chemotactic to myeloid cells, and may also have a role in cell migration, activation, and chemotaxis (7). Although EPCs are rare in the circulation, they can be mobilized into circulation from bone marrow by vascular trauma or systemic administration of cytokines (12, 13, 15, 16). Multipotent adult progenitor cells (MAPC) derived from postnatal human bone marrow were demonstrated to be progenitors for angioblasts and to undergo a differentiation process from CD34(−), VE-cadherin(−) cells to CD34(+), VE-cadherin(+) cells after culture in the presence of VEGF. These cells subsequently differentiate into morphologically and functionally mature endothelial cells that contribute to neoangiogenesis in vivo during tumor angiogenesis and wound healing (15,17).
The role of HIMF in hypoxic tissue and how HIMF exerts its angiogenic and vasoconstrictive properties remain unclear. The current work defines a targeted molecular binding partner of HIMF and investigates whether HIMF functions as a chemotactic molecule for recruiting bone-marrow derived cells to hypoxic or ischemic tissue like its homolog XCP1. To determine HIMF's major binding partner(s) we used GST-pull-down and mass spectrometry techniques. We isolated a HIMF-binding molecule, BTK, a molecule known to be crucial in regulation of B-cell maturation and involved in cell migration. BTK stimulates B cell differentiation in bone marrow, and mutations in BTK are responsible for X-linked agammaglobulinemia (XLA) in humans and X-linked immunodeficiency (xid) in mice (18, 19). Since HIMF expression is induced in hypoxic tissue, and bone marrow derived EPCs are preferentially recruited to the site of ischemic tissue, we investigated whether HIMF is a chemotactic molecule for bone marrow cells.
There is a continuing need in the art to identify important targets for treating pulmonary, cardiac, and inflammatory diseases.